gastrin-17 and Pancreatic-Neoplasms

Aphton is developing an anti-gastrin vaccine [Anti-gastrin 17 immunogen, G17DT, Gastrimmune], which neutralises the gastrin 17 hormone and the Gly-G17 hormone. Gastrin 17 is a growth factor for pancreatic, stomach and colorectal cancers, and a potent stimulator of gastric acid secretion.The anti-gastrin immunogen, G17DT, consists of a large carrier protein, called Diptheria Toxoid (DT). A synthetic peptide, which is similar to a portion of the gastrin 17 hormone (GT), is attached to the carrier protein. These are then contained in a liquid suspension vehicle. When administered to patients, G17DT induces an immune response producing antibodies, which cross-react and neutralise the target hormone thus preventing its interaction with disease-causing or -participating cells. Aphton has entered into an agreement with Aventis Pasteur for the marketing of G17DT in all human cancer indications in North America and Europe. Under the terms of the agreement, Aphton is responsible for product development, clinical trials and regulatory agency approvals. The agreement was originally with Pasteur Mérieux Connaught, a subsidiary of Rhône-Poulenc. However, in December 1999, Rhône-Poulenc merged with Hoechst to form Aventis. As a result of the merger, Pasteur Mérieux Connaught underwent a name change to Aventis Pasteur. In July 2002, Aphton announced that it would negotiate with companies wanting to licence the vaccine in territories other than North America and Europe. In February 2003, Aphton announced it had received fast track designation for G17DT in combination with cisplatin and fluorouracil for use in stage IV gastric cancer to improve overall survival. In July 2002, the US FDA granted G17DT orphan drug status for the treatment of gastric cancer, an indication that was broader than the indication Aphton originally sought. The vaccine was also granted orphan drug status in Australia for gastric cancer in December 2002. In July 2002, Aphton announced that the US FDA had granted G17DT orphan drug status for the treatment of adenocarcinoma of the pancreas. In September 2002, the US FDA also granted G17DT, used in combination with gemcitabine, fast track status for the treatment of pancreatic cancer patients. In addition, the vaccine was also granted orphan drug status in Australia for pancreatic cancer in December 2002. In March 2003, Aphton announced that the Committee for Orphan Medicinal Products had recommended to the European Commission that G17DT be granted orph

Topics: Animals; Cancer Vaccines; Clinical Trials as Topic; Colorectal Neoplasms; Gastrins; Humans; Orphan Drug Production; Pancreatic Neoplasms; Peptic Ulcer; Rats; Stomach Neoplasms; Vaccines

This study aimed to investigate G17DT, an immunogen producing neutralizing antibodies against the tumor growth factors amidated and glycine-extended forms of gastrin-17, in the treatment of pancreatic cancer.. A randomized, double-blind, placebo-controlled, group-sequential multicenter trial of G17DT in patients with advanced pancreatic cancer unsuitable for or unwilling to take chemotherapy. Inclusion criteria were a Karnofsky performance score of 60 or higher and a life expectancy of more than 2 months. Patients received G17DT or placebo emulsion at weeks 0, 1, 3, 24, and 52. The primary end point was survival, and secondary end points were tolerability, Karnofsky performance.. A total of 154 patients were recruited: 79 G17DT and 75 placebo. A final analysis of the intention-to-treat population, using a proportional hazards model, stratifying by disease stage and adjusting for interim analysis, gave a hazard ratio for mortality of 0.75 (95% confidence interval, 0.51-1.10, P = 0.138; G17DT/placebo). A conventional analysis without adjustment for disease stage or interim analysis, censoring for chemotherapy and excluding protocol violators, gave median survival periods of 151 (G17DT) and 82 days (placebo) (log-rank test, P = 0.03).Patients developing anti-G17DT responses (73.8%) survived longer than nonresponders or those on placebo (median survival, 176 vs 63 vs 83; log-rank test, P = 0.003). G17DT was well tolerated.

Topics: Adult; Aged; Aged, 80 and over; Cancer Vaccines; Double-Blind Method; Europe; Female; Gastrins; Humans; Kaplan-Meier Estimate; Karnofsky Performance Status; Life Expectancy; Male; Middle Aged; Pancreatic Neoplasms; Placebos; Proportional Hazards Models; Prospective Studies; Time Factors; Treatment Outcome

Two major forms of gastrin, gastrin-17 (G17) and gastrin-34 (G34), exist in blood. However, conventional immunoassay methods can only quantify total gastrin or G17 alone. Here, we aimed to establish a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to quantify G17 and G34 simultaneously.. Serum samples were prepared by anion-exchange solid-phase extraction. The analytical performance of the LC-MS/MS method was validated and the method was compared to chemiluminescence immunoassay (CLIA) and radioimmunoassay (RIA). The G17 and G34 concentrations in 245 serum samples from healthy controls, individuals with gastrinoma, and individuals with other diseases were analyzed.. The total runtime of the LC-MS/MS method was 6 min. No substantial matrix effect was observed with internal standard correction. The intraassay coefficients of variation (CVs) for G17 and G34 were 4.0%-14.2% and 4.4%-10.4%, respectively, and total CVs were 5.2%-14.1% and 4.6%-12.4%, respectively. The correlation coefficient between LC-MS/MS and CLIA was 0.87, and between LC-MS/MS and RIA was 0.84. The G17+G34 concentrations for 87.5% of individuals with gastrinoma were higher than the 95th percentile of healthy controls (18.1 pg/mL), whereas the concentrations for individuals with other diseases and gastrinoma overlapped. Based on the Youden indices calculated for G17+G34, G34, and G17, the most specific biomarker was G17 (96.9% clinical specificity at 209.8 pg/mL) for gastrinoma.. This method should aid in the diagnosis of diseases associated with increased gastrin concentrations.

Topics: Chromatography, Liquid; Gastrinoma; Gastrins; Humans; Pancreatic Neoplasms; Tandem Mass Spectrometry

The aim of the study was to evaluate the anti-tumor mechanism of Z-360, a gastrin/cholecystokinin-2 receptor (CCK2R) antagonist, in MIA PaCa-2 cells and in a subcutaneous xenograft mice model.. The anti-tumor effects of Z-360 and/or gemcitabine were monitored using a MIA PaCa-2 xenograft model. The effect of Z-360 on apoptosis in the model was examined by TUNEL staining and real-time PCR analysis and the effect in MIA PaCa-2 cells stably expressing human CCK2R was also evaluated by caspase-3/7 activity.. In this xenograft model, Z-360 significantly reduced the tumor weight, increased TUNEL-positive cells and suppressed the expression of anti-apoptosis factors such as survivin, XIAP and Mcl-1, and these effects of Z-360 combined with gemcitabine were more effective. Furthermore, gastrin-17 and gastrin-34 inhibited apoptosis in vitro and Z-360 dose-dependently abrogated this effect.. These results suggest that Z-360 exerts an anti-tumor effect through a reduction in anti-apoptosis factors by blocking CCK2R.

Topics: Animals; Apoptosis; Benzodiazepinones; Cell Line, Tumor; Cell Proliferation; Deoxycytidine; Endopeptidases; Gastrins; Gemcitabine; Gene Expression Regulation, Neoplastic; Humans; Inhibitor of Apoptosis Proteins; Mice; Myeloid Cell Leukemia Sequence 1 Protein; Pancreatic Neoplasms; Receptor, Cholecystokinin B; Survivin; X-Linked Inhibitor of Apoptosis Protein; Xenograft Model Antitumor Assays

Gastrin is known to enhance the growth of pancreatic carcinoma via the cholecystokinin (CCK)-2/gastrin receptor. We investigated the anti-tumor effect of Z-360 (calcium bis [(R)-(-)-3-[3-{5-cyclohexyl-1-(3,3-dimethyl-2-oxo-butyl)-2-oxo-2,3,4,5-tetrahydro-1H-benzo[b][1,4]diazepin-3-yl}ureido]benzoate]), a novel orally active CCK-2 receptor antagonist alone or combined with the chemotherapeutic agent, gemcitabine in human pancreatic adenocarcinoma cell lines.. Z-360 potently inhibited specific binding of [3H]CCK-8 to the human CCK-2 receptor, with a Ki value of 0.47 nmol/l, and showed antagonistic activity for this receptor. The anti-tumor effect of Z-360 alone or combined with gemcitabine was assessed using subcutaneous xenografts of MiaPaCa2 and PANC-1 and an orthotopic xenograft model (PANC-1). Oral administration of Z-360 significantly inhibited the growth of MiaPaCa2 (41.7% inhibition at 100 mg/kg, P<0.01). Combined administration of Z-360 and gemcitabine significantly inhibited subcutaneous PANC-1 tumor growth compared with either agent alone (27.1% inhibition compared to effect with gemcitabine, P<0.05), and significantly prolonged survival compared with the vehicle control (median survival of 49 days in vehicle compared to 57 days in the combination group, P<0.05). In vitro studies showed that Z-360 significantly inhibited gastrin-induced proliferation of human CCK-2 receptor-expressing cells, and also significantly reduced gastrin-induced PKB/Akt phosphorylation to the level of untreated controls.. In the present study, we have shown that Z-360 combined with gemcitabine can inhibit pancreatic tumor growth and prolong survival in a pancreatic carcinoma xenograft model, on a possible mode of action being the inhibition of gastrin-induced PKB/Akt phosphorylation through blockade of the CCK-2 receptor. Our results suggest that Z-360 may be a useful adjunct to gemcitabine for the treatment of pancreatic carcinoma and a therapeutic option for patients with advanced pancreatic cancer.

Topics: Adenocarcinoma; Administration, Oral; Animals; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Benzodiazepinones; Cell Line, Tumor; Cell Proliferation; Deoxycytidine; Disease Models, Animal; Female; Gastrins; Gemcitabine; Humans; Mice; Mice, Nude; Pancreatic Neoplasms; Phosphorylation; Proto-Oncogene Proteins c-akt; Receptor, Cholecystokinin B; Survival Rate; Xenograft Model Antitumor Assays

The interaction of gastrin with the cholecystokinin 2 (CCK2)/gastrin receptor has been studied extensively in relation to gastric acid secretion. However, not much is known about the contribution of individual amino acids of gastrin interacting with the CCK2 receptor, when gastrin is acting as a tumor growth factor. The purpose of the present study was to determine the significance of each individual amino acid residue of human gastrin-17 with respect to CCK2 receptor-mediated cell proliferation. Activation of this receptor was assessed using an in vitro bioassay based on gastrin-induced expression of a c-fos-luciferase reporter, transfected in AR42JB13 and Colo 320 cells, a rat pancreatic and human colorectal cell line respectively. Gastrin-17 dose dependently increased c-fos induction in both cancer cell lines. L365,260, a known CCK2 receptor antagonist, completely blocked the gastrin signal, demonstrating the specificity of this assay. We demonstrated for the first time that four carboxy-terminal amino acids of gastrin-17 are essential for activation of the CCK2 receptor with respect to c-fos induction. Also other residues of gastrin-17, notably glycine-2 for the rat CCK2 receptor and glutamic acid 8-10 and tyrosine-12 for the human receptor, were found to be important, although to a lesser extent. Alanine-substitution variants of each of the four carboxy-terminal amino acids of gastrin-17 showed strongly reduced receptor activation but did not act as competitive inhibitors of gastrin-17. Identification of the essential role of the carboxy-terminal tetrapeptide of gastrin-17 in CCK2 receptor-mediated c-fos induction indicates that gastrin inhibitory therapeutic strategies should mainly be targeted toward this region of gastrin.

Topics: Alanine; Amino Acid Substitution; Animals; Cell Proliferation; Colorectal Neoplasms; DNA Primers; Gastrins; Genes, fos; Humans; Luciferases; Pancreatic Neoplasms; Pentagastrin; Promoter Regions, Genetic; Protein Precursors; Rats; Receptor, Cholecystokinin B; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tumor Cells, Cultured

To investigate the effects of gastrin and cholecystokinin (CCK) and their specific antagonists on the growth of pancreatic and biliary tract cancer cell lines.. Five pancreatic and 6 biliary cancer cell lines with 2 conrtol cells were used in this study. Cell proliferation study was done using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) test and direct cell count method. Reverse transcription-polymerase chain reaction (RT-PCR) and slot blot hybridization were performed to examine and quantify the expression of hormonal receptors in these cell lines.. SNU-308 showed a growth stimulating effect by gastrin-17, as did SNU-478 by both gastrin-17 and CCK-8. The trophic effect of these two hormones was completely blocked by specific antagonists (L-365, 260 for gastrin and L-364, 718 for CCK). Other cell lines did not respond to gastrin or CCK. In RT-PCR, the presence of CCK-A receptor and CCK-B/gastrin receptor mRNA was detected in all biliary and pancreatic cancer cell lines. In slot blot hybridization, compared to the cell lines which did not respond to hormones, those that responded to hormones showed high expression of receptor mRNA.. Gastrin and CCK exert a trophic action on some of the biliary tract cancers.

Topics: Biliary Tract Neoplasms; Cell Count; Cell Division; Cell Line, Tumor; Cholecystokinin; Gastrins; Hormone Antagonists; Humans; Pancreatic Neoplasms; Receptor, Cholecystokinin A; Receptor, Cholecystokinin B; Reverse Transcriptase Polymerase Chain Reaction; Tetrazolium Salts; Thiazoles

Topics: Adult; Antibodies; Enzyme-Linked Immunosorbent Assay; Female; Gastrin-Secreting Cells; Gastrinoma; Gastrins; Humans; Male; Pancreatic Neoplasms; Radioimmunoassay

Functional gastrin-containing tumor cells were maintained for up to 8 wk without fibroblastoid cell overgrowth. Short-term cultures consisted mainly of colonies composed of small polygonal cells, 70%-90% of which stained positive for immunoreactive gastrin. Cultures exhibited limited growth but viability remained high for 2-3 wk. Culture medium contained component I, and gastrin 34, 17, and 14. With time the major C-terminal gastrin species in medium changed from gastrin 17 at 3 days to gastrin 34 at 5 wk. Extracts of cultured cells contained gastrin 34, 17, and 14; gastrin 17 was the major form detected at all times. Ultrastructurally, cultured tumor cells retained morphological integrity for several weeks; however, with time changes in the appearance of the secretory granules accompanied by evidence of cellular retrodifferentiation were gradually observed. Secretin, gastrin-releasing peptide, 8-bromoadenosine 3':5'-cyclic monophosphate, and phorbol, 12-myristate, 13-acetate stimulated the release of gastrin from cultured cells in a time-dependent fashion. Secretin, bombesin, gastrin-releasing peptide, L-tryptophan, and ethylamine stimulated gastrin release in a dose-dependent fashion. Somatostatin 14 inhibited secretin, bombesin, and gastrin-releasing peptide stimulated gastrin release but did not alter basal release. Cultured cells demonstrated de novo gastrin synthesis, evidenced by their ability to incorporate radiolabeled amino acids into immunoadsorbable gastrinlike material. Primary cultures of gastrin-containing tumor cells free from stromal contamination offer unique advantages for studies of factors that regulate the synthesis and secretion of gastrin and may prove of potential value for studies on cell differentiation and growth.

Topics: Culture Media; Gastrinoma; Gastrins; Humans; Immunoenzyme Techniques; Pancreatic Neoplasms; Protein Precursors; Time Factors; Tumor Cells, Cultured

Tissue pieces of a metastatic human gastrinoma (ultrastructural Type II) were successfully transplanted to the anterior eye-chamber of rats immunosuppressed with Cyclosporin A. Immunocytochemical investigation of the transplants showed evidence for preserved endocrine activity of tumour cells with immunoreactivity towards the C-terminal of the gastrin/cholecystokinin molecule. Studies of gastric acid secretion in tumour-bearing rats and sham-operated controls with chronic gastric fistulas showed that the basal acid output did not differ between the groups during 3 weeks of study. However, the stimulated gastric acid secretion decreased after 5 days in both groups to remain significantly depressed throughout the study, an effect probably due to Cyclosporin A treatment of the groups. The concentration of immunoreactive gastrin in plasma from rats with tumours in oculo was 5 times higher than in sham-operated rats. Gastrin-34 was the major immunoreactive component in both patient serum and rat plasma. An immunoreactive fraction corresponding to component I was found in the patient serum, but not in the rat plasma, although present in the chamber fluid. Components corresponding to gastrin-17 were found both in the patient serum and in the rat plasma. The chromatographic pattern of the tumour was similar to that in rat chamber fluid. The dominating component corresponded to gastrin-17, while gastrin-34 represented the quantitatively smaller component. Gastrin-34 was, however, relatively more abundant in the tumour extract than in the chamber fluid. The study also indicates that a gastrin-producing tumour transplanted in oculo in immunosuppressed rats may increase the rat plasma concentration of the same molecular forms of gastrin as seen in the clinical situation.

Topics: Adolescent; Animals; Anterior Chamber; Chromatography, Gel; Female; Gastric Acid; Gastrinoma; Gastrins; Humans; Immunosuppression Therapy; Male; Microscopy, Electron; Neoplasm Transplantation; Pancreatic Neoplasms; Protein Precursors; Radioimmunoassay; Rats; Rats, Inbred Strains